published_or_final_version / Medicine / Doctoral / Doctor of Philosophy

<p>Obesity is today a problem that has reached epidemic proportions. One of the causes of obesity is the over-consumption of energy. Fat is the most energy-dense nutrient, where the quality seems to be more important for the development of the metabolic diseases than the quantity. The fatty acid composition in serum lipid fractions can be used to mirror the dietary fat quality.</p><p>Stearoyl-CoA-desaturase (SCD) is an enzyme that converts saturated to monounsaturated fatty acids. A surrogate measure of SCD activity can be estimated as a fatty acid ratio; 16:1/16:0 (palmitoleic acid/palmitic acid) and 18:1/18:0 (oleic acid/stearic acid). The aim of this project was to investigate the intra-day variation in the SCD-ratio in humans eating a standardized diet. The results showed that triacylglycerol and nonesterified fatty acid fractions in serum lipids had a significant variance in the 16:1/16:0 ratio during the day, whereas 18:1/18:0 ratio in the same fractions did not exhibit the same pattern. In this study 16:1/16:0 ratio also seems to be a better marker than 18:1/18:0 ratio for estimating SCD activity. For further evaluation of the intra-day variation there need to be a more long-term study of the SCD-activity for a larger group of subjects.</p>

Three studies were designed to evaluate the promoting effect of cyclopropenoid fatty acids (CPFA) on avian myeloblastosis virus (AMV) infection in White Leghorn chickens. In the first study effects on growth parameters, blood serum and erythrocyte lipid composition of White Leghorn chickens were investigated at 100 ppm and 300 ppm dietary CPFA. In comparison to controls, chicks fed dietary CPFA had reduced body weights and food intake. At both levels, CPFA increased the saturated fatty acids and decreased monoenes in blood serum and erythrocyte triacylglycerols and phospholipids. In a second study, the effects of high dietary (300 ppm) CPFA on AMV infectivity were investigated. Chicks were inoculated with 0.1 ml AMV at 7 days of age. A diet containing CPFA was fed before and after inoculation. CPFA/AMV treated chicks showed a marked reduction in body weight gain and food consumption in comparison to controls. CPFA appeared to promote AMV infection along with unexpected violent hemorrhagic manifestations. These hemorrhagic manifestations appeared to cause death before AMV induced myeloblast proliferation in the circulating blood. In comparison to controls, CPFA altered the phospholipid composition of isolated myeloblasts and the cholesterol/phospholipid ratio of isolated myelobasts, myeloblast plasma membrane, and virus particles. Analysis of the acyl group composition of choline and ethanolamine phospholipids from CPFA treated myeloblasts and myeloblast plasma membrane revealed an increase in saturated fatty acid and a decrease in monoenes. In both, CPFA appeared to affect ethanolamine more than choline phospholipids. In contrast, the alterations observed in CPFA treated virus particle choline and ethanolamine fatty acyl chains were reversed from those found in the myeloblast and myeloblast plasma membrane. In addition, ATPase specific activity was altered in relation to the CPFA induced lipid changes in the myeloblast plasma membrane and virus particle. It was shown that as saturation increased enzyme activity increased. As a third investigation, the effects of low dietary (75 ppm and 150 ppm) CPFA on AMV infection was conducted in an attempt to eliminate the hemorrhagic manifestations. A difference that was observed between this investigation and the 300 ppm CPFA/AMV study was less severe hemorrhagic responses. Histological examination of CPFA treated liver and spleen by light microscopy revealed heavy infiltration of mitotic myeloblasts into each tissue. Analysis of the number of virus particle/ml by reverse transcriptase revealed 300 ppm dietary CPFA stimulated the synthesis and release of virus particles. This was in contrast with 150 ppm dietary CPFA which retarded the synthesis and release of virus particles. / Graduation date: 1982

The effect of vitamin B-6 (B-6) status on plasma fatty acids (FA) levels and lipid metabolism was investigated in this metabolic study. Eight female subjects were fed for 28 days. For the first 7 days, they were fed a constant diet containing 2.10 mg of B-6. For the rest of the period (21 days), they were differentiated in terms of B-6 intake; 4 of them were fed a low (0.93 mg/day) and 4 a high (2.60 mg/day) B-6 diet. B-6 status indices, plasma FA concentration and lipid profile were determined. Plasma pyridoxal 5'-phosphate and total B-6 concentration (P<0.01), urinary 4- pyridoxic acid and total B-6 concentration (P<0.001) showed a significant difference between the two groups at the end of the study. Erythrocyte PLP failed to show any significant difference between the two groups throughout the diet study. There was no significant difference in the plasma FA or lipid profile between the two groups. Plasma total cholesterol (TC) of the low B-6 group decreased slightly (7 %), but was not statistically significant. When comparing day 7 and day 28 values, plasma triglycerides increased (9 %) for the high and decreased for the low B-6 group. LDL-C decreased (5 %) for the high B-6 group but did not change in the low B-6 group. HDL-C decreased slightly in both groups (~8 %). There was no clear evidence that a low intake of vitamin B-6 affects the fatty acid and lipid metabolism. Further studies are required to identify the relationship between vitamin B-6 and fatty acid and lipid metabolism in humans. / Graduation date: 1997

Cyclopropenoid fatty acids (CPFA), which are a group of fatty acids produced by plants of the order Malvales, are known to induce adverse physiological effects when administered to a variety of animal species. A structurally strained cyclopropene ring is present in all CPFA and is believed responsible for the toxic action of these fatty acids. Dietary consumption of CPFA by mammals, poultry and fish has resulted in toxic responses including hepatic damage, impaired reproductive capabilities and sizeable alterations in lipid metabolism. Furthermore, CPFA have been identified as mildly carcinogenic and strongly cocarcinogenic towards rainbow trout (Salmo gairdneri). The mechanism by which CPFA enhance carcinogenesis is currently not understood. The research in this thesis has therefore been directed toward obtaining a better understanding as to how CPFA induce toxic responses in rainbow trout. Hepatic plasma membranes were isolated from both control trout and trout which had consumed dietary CPFA. The plasma membranes were then compared via the use of electron microscopy, chromatographic analysis of phospholipid and fatty acid content, two dimensional polyacrylamide gel electrophoresis of proteins, and Western blot analysis of concanavalin A sensitive glycoproteins. Electron micrographs revealed that control plasma membranes appeared more homogeneous than CPFA membranes and were characterized by more membrane sheets and less vesicularization. The analysis of enzyme activities revealed that CPFA caused a decrease in whole liver glucose-6-phosphatase activity and that control plasma membranes expressed slightly higher glucose-6-phosphatase and 5'-nucleotidase activities as compared to CPFA membranes. Although dietary CPFA appeared to have no effect on the phospholipid content of the plasma membranes, significant alterations in the fatty acid profiles of ethanolamine and choline phospholipids were observed. CPFA caused a decrease in palmitic, palmitoleic and oleic acids while the level of stearic and docosahexaenoic acids subsequently increased. Differences between the protein content of control and CPFA plasma membranes were made clear through the analysis of electrophoretic and Western blotting data. Membranes isolated from fish fed CPFA contained several proteins of high molecular weight (above 66,000 daltons) and other proteins of high isoelectric point that were not present in control plasma membranes. Additionally, two families of glycoproteins which had previously been identified as microsomal in origin were detected only in CPFA plasma membranes. A discussion concerning the possible causes and biological ramifications of the observed subcellular alterations caused by CPFA insult is also presented in this thesis. / Graduation date: 1989

The effect of different blanching treatments and packaging materials on the enzymatic (lipoxygenase and peroxidase) activity and fatty acid stability of two different varieties of sweet corn on the cob (Jubilee and GH 2684) was evaluated during nine months of frozen storage at -23.3°C. The initial moisture content in the kernels of the two sweet corn varieties averaged 72.5%. After nine months of frozen storage the moisture content in the kernels of corn depended greatly on the packaging material used. The ears stored in Cryovac B and E bags showed the best moisture retention (72.2% final moisture content), followed by the polyethylene bags (71.4%) while the ears stored without packaging material showed severe dehydration (70.1%). The peroxidase and lipoxygenase activities were determined using spectrophotometric assays on a crude extract obtained from liquid nitrogen powdered corn. Both unblanched varieties of sweet corn showed similar initial peroxidase specific activity and general behavior during the nine months of frozen storage. The presence of lipoxygenase isozymes with different thermal stabilities in both varieties was suggested by the higher lipoxygenase specific activity found in Jubilee after freezing and nine months of frozen storage (0.135 units/mg protein) compared with the GH 2684 variety (0.115 units/mg protein). Complete inactivation of lipoxygenase was obtained after 9 minutes steam blanching at 100°C. Peroxidase was more heat resistant showing some remaining specific activity after 9 minutes steam blanching with a complete inactivation after 15 minutes steam blanching. No regeneration of either enzyme was observed during the nine months of frozen storage suggesting a permanent disruption of the active site of both enzymes. Relative fatty acid content was determined by gas chromatographic analysis of fatty acids methyl esters. The major fatty acids present in both varieties were palmitic (14.93%), stearic (2.79%), oleic (31.54%), linoleic (46.87%) and linolenic (1.89%) acids. Good stability of the polyunsaturated fatty acids was observed during the nine months storage at -23.3°C, with autoxidation as the main mechanism responsible for the decrease in the relative percent of polyunsaturated fatty acids. Some enzymatic oxidation also occurred, decreasing the linolenic acid content. The control of the degradation of polyunsaturated fatty acids depended mostly on the frozen storage temperature (-23.3°C) and not on the oxygen permeability of the different packaging materials. The results obtained in our study suggested that blanching of the ears of sweet corn had an important effect on reducing the enzyme activity but little effect on the polyunsaturated fatty acid degradation after 9 months of storage at -23.3°C. / Graduation date: 1994

The DNA repair capabilities of rainbow trout (Salmo gairdneri) were studied vising the method of autoradiography. Trout were fed a semi-purified control diet containing 0 ppm, 50 ppm, or 300 ppm cyclopropenoid fatty acids (CPFA) for 6-9 weeks. Liver slices were prepared and exposed in vitro to a control treatment, ultraviolet irradiation (UV), ethidium bromide (EB), UV/EB in succession, or aflatoxin B₁. The degree of DNA repair was analyzed in terms of net grains per cell. Except following the EB treatment, fish on the control diet revealed an absence of ongoing DNA repair. Trout fed 50 ppm CPFA exhibited a consistently low level of repair over time following the in vitro control treatment. Fish fed 300 ppm CPFA revealed a relatively higher degree of ³H-Me-thymidine incorporation indicative of induced DNA repair following the in vitro control treatment, and the degree of repair increased with time on the diet. UV-irradiation caused a marked increase in the degree of induced DNA repair in 300 ppm CPFA fish at 6 and 7.5 weeks, and in 50 ppm CPFA fish at 7.5 weeks. Follcwing UV-irradiation, liver slices were exposed to EB, a DNA intercalating agent used to inhibit normal DNA replication. However, in contrast to the desired effect, EB caused a marked decrease in the degree of repair synthesis observed in 300 ppm CPFA fish at 6 and 7.5 weeks. Indicative of intercalation, the in vitro EB treatment caused a moderate degree of ³H-Me-thymidine incorporation in fish fed the control diet. Repair was also induced in 300 ppm CPFA fish following exposure to EB at 6 and 7.5 weeks. Aflatoxin B₁ induced DNA repair to various degrees in fish on all diets at 7.5 and 9 weeks. In comparison to the in vitro control treatment, it was observed that the degree of induced DNA repair was decreased significantly - "completely" following the UV, UV/EB, and EB treatments - in fish fed the 300 ppm CPFA diet for 9 weeks. In view of the low level of DNA repair observed in rainbow trout using autoradiography, the repair capabilities were studied using a more sensitive assay, bromodeoxyuridine (BrdU) photolysis. Isolated hepatocytes were prepared from fish fed the various diets and exposed in vitro to a control treatment, UV-irradiation, or 4-nitroquinoline-N-oxide. The obtained results were nonconclusive indicating technical improvements on the assay need to be made. / Graduation date: 1988

No description available.

The present research examined the hypothesis that omega-3 polyunsaturated fatty acids would increase seizure threshold in rats in vivo, and reduce neuronal excitability in mouse hippocampal slices. Seizure thresholds were measured in rats using the maximal pentylenetetrazol and electrical stimulation seizure tests following α-linolenic acid (ALA) or docosahexaenoic acid administration. ALA raised seizure threshold in the maximal PTZ seizure test, but this effect probably occurred because ALA displaced DHA from liver to the brain. DHA itself was therefore tested in the PTZ and electrical stimulation seizure tests. Direct administration of DHA by subcutaneous injection raised seizure thresholds in the PTZ seizure test, which models tonic-clonic attacks in humans. Dietary enrichment with DHA raised afterdischarge seizure thresholds in the cortex and amygdala, which model simplex and complex partial seizures in humans, although this effect took some time to occur. In vitro, the application of DHA also reduced the incidence of excitatory sharp waves in mouse hippocampal slices. This effect did not appear to be due to either an increase in GABAergic inhibitory tone, nor to a decrease in glutamatergic drive. The fatty acid composition of phospholipids and unesterified fatty acids were measured in the brain following microwave fixation in order to determine whether the effects of DHA on seizure thresholds were due to its de-esterification from the phospholipid membrane. The assay surprisingly revealed that subcutaneous administration of DHA at a dose that raised seizure threshold, increased unesterified arachidonic acid, but not unesterified DHA concentrations during seizures. The results of these studies support the hypothesis that DHA raises seizure threshold in rats, and reduces neuronal excitability in vitro. The effects of DHA on seizure threshold are possibly mediated by the de-esterification of arachidonic acid, which is known to have effects on the voltage-dependent sodium channel.

Resveratrol (RSV) is a natural compound that improves mitochondrial function and metabolic health in animal models. Thus far, RSV’s effects on metabolic outcomes in humans are controversial, and RSV’s acute mechanism has not yet been confirmed in vivo. This study aimed to evaluate the effect of an acute dose of RSV on insulin sensitivity and fatty acid oxidation, and to determine RSV’s mechanism of action in human skeletal muscle and adipose tissue. Overweight males (n=8; BMI, 30.5±3.6; VO2peak, 34.0±7.3 ml/kg) reported to the lab on 2 occasions and were provided a breakfast supplemented with 0.3g of RSV or a placebo pill. Experiments were performed in random order using a double blind crossover design. Gas exchange measures, blood samples, and skeletal muscle and adipose tissue biopsies were obtained before and 2 hours after the supplement meal. RSV acutely improved insulin sensitivity, but had no effect on fatty acid oxidation. Additionally, RSV supplementation had no effect on the intracellular signaling of key proteins proposed to mediate its effects in skeletal muscle and adipose tissue. Taken together, these results suggest a single dose of RSV can acutely enhance insulin sensitivity, but its mechanism of action is not conserved across species, and its intracellular signaling pathway is different in humans than previously thought. Due to its insulin sensitizing effect, RSV retains its clinical value, but further research is required to determine its most useful application for human metabolic health. / Thesis (Master, Kinesiology & Health Studies) -- Queen's University, 2013-06-06 13:30:03.522